The protection of biologically and/or pharmaceutically active substances like drugs and especially proteins intended to be applied for medical uses against denaturation and enzymatic degradation is an important issue for all drug delivery systems. Also the controlled release of such active compounds, including small molecules, after application and the enhancement of transport across mucosal surfaces remain important issues.
Polymeric excipients as possible drug delivery systems are in principle known in the state of the art. Possible strategies based on polymeric carriers for mucosal and parenteral delivery of proteins include for example (a) modification of biologically active compounds with polymers, (b) encapsulation of the hydrophilic macromolecules into micro- or nanospheres or micro- or nanoparticles, and (c) adsorptive drug loading onto the surface of micro- or nanospheres or micro- or nanoparticles. Because of their medical use these systems are also required to be bio-degradable.
This challenge is especially important for oral, mucosal and parenteral systems designed for hydrophilic, especially macromolecular ingredients. Additionally, each requirement differs with regard to the specific envisaged application, which motivates a search for chemically diverse polymers.
Some polymers have already been developed to meet these requirements, but there still remains a formidable challenge since structure function relations of suitable polymers are still scarce. A certain approach aims at the identification of negatively charged polymers, especially polyesters as carriers for pharmacologically active substances because these are expected to be particularly suitable for effective loading and to exhibit sustained-release properties of positively charged drugs, proteins and peptides.
Biodegradable matrix polymers for active ingredient embedding have already been described in U.S. Pat. No. 3,773,919. Polymers from hydroxycarboxylic acids, especially lactic and/or glycolic acid were proposed. However, depot forms from polylactic acid (PLA) or polylactic-co-glycolic acid (PLGA), especially microparticles, generally exhibit a multiphase release trend and initially display a sharply increased release because of active ingredient present on the surface. This is followed by a phase of sharply reduced or nonexistent release, especially in peptide active ingredients, which is then followed by later active ingredient liberation supported by polymer mass degradation. Polymer residues are still present at the time of completion of active ingredient release.
EP 058481 A1 describes the use of a mixture of PLGA having different molecular weights. Drug release is supposed to be linearized by this and the degradation rate adjusted to the release period. Use of such polymer mixtures, however, imposes high requirements on the hydrolytic stability of the active ingredient and they are generally not well suited for the production of microparticles.
DE 3430852 A1 describes esters from polyols and poly- or copolylactic acid, which are also proposed as matrix polymers for depot forms. Example 26 entails in-vitro release of washed microparticles containing bromocriptin mesylate produced by spray drying. Despite washout of the active ingredient adhering to the microparticle surface, after 24 hours 62% of the active ingredient load had already been released.
EP 407617 A1 describes a biocompatible polyester with increased hydrolysis rate consisting of saccharides bonded to PLA or PLGA. The polyesters are proposed as matrix material for depot forms. However, they seem to be associated with the problem of nonuniform active ingredient release and especially the problem of the initial burst effect.
Additionally, polyesters consisting of a polysaccharide backbone are thermally not very stable during the grafting reaction, and are also less soluble in monomer melt as compared to polyesters with a polyvinyl backbone. In addition, the presence of the naturally occurring saccharide may imply the risk of possible immunogenic reactions in vivo. Furthermore, these polymers show bulk erosion.
DE 19839515 A1 discloses the colloidal association of an active agent like a peptide, a DNA construct or a vaccine with a polyol ester which together in the form of a colloid are to be used as a pharmaceutical preparation for controlled transmucosal administration. The disclosed polymers for such a use are branched polyol esters consisting of a central molecule to which short-chain, biodegradable hydroxycarboxylic acid ester groups are attached.
Polymers, based on a polyvinyl alcohol backbone are described in the publication “Biodegradable comb polyesters: Part 1; synthesis, characterization and structural analysis of poly(lactide) and poly(lactide-co-glycolide) grafted onto water-soluble poly(vinyl alcohol) as backbone” by Breitenbach et Kissel, Polymer, 39 (14): 3261-3271 (1998). Based on an analysis of their physicochemical properties they are described as having considerable potential as a parenteral drug delivery system for peptides and proteins.
Patent application EP 1132416 A1 discloses colloidal nanoparticular carriers comprising loaded or non-loaded water soluble comb polymers and their use in mucosal applications. They are characterized by a backbone formed from water-soluble polyol(s) grafted with hydrophobic side-chains, providing an amphiphilic character, and optionally ionic groups, where the backbone polymer has a weight average molecular weight (Mw) of 10,000-30,000 g/mol and the side-chains preferably have a combined Mw of 45,000-100,000 g/mol. Such colloidal nanoparticular carriers are described as being useful as drug delivery systems for especially large biomolecules like proteins and nucleic acids. Unfortunately, because of their ability to attach to mucous membranes (bio-adhesion) they are expected to induce a systemic immune response.
US patent application US 2001/0047074 A1 (granted as U.S. Pat. No. 6,616,944 B2) discloses self-assembling, polymer-based delivery systems for proteins. The delivery systems comprise an active agent and a polyol ester, having a linear polyol containing six or more hydroxyl groups as a central backbone and biodegradable hydroxy carboxylic ester groups attached to the central backbone; as an additional feature of these systems, the linear polyol contains charged groups, proton donating groups, and/or proton accepting groups, which are attached via a spacer group or an ether-, ester-, or urethane-linkage to the linear polyol. Such delivery systems are described to form stable complexes with proteins and therefore as being useful as drug carriers for therapeutic use or vaccines.
Unfortunately, co-polyesters of just lactic and glycolic acid (PLGA) turned out to be suboptimal for protein and DNA delivery due to inactivation by the acidic microenvironment and uncontrolled burst release due to poor compatibility between lipophilic polymers and hydrophilic drug candidates.
One proposed solution to address these problems associated with PLGA has been to introduce the hydrophilic composition with functional group to this polyester system, leading to first copolymers of PLGA and poly(ethylene glycol)(PEG), as described e.g. in: Youxin, L., Kissel T.: Synthesis and properties of biodegradable ABA triblock copolymers consisting of poly(L-lactic acid) or poly(L-lacticgo-glycolic acid) A-blocks attached to central poly(oxyethylene) B-blocks; J. Control Release 1993; vol. 27, pages 247-257.
Subsequently described approaches have been:                the development of star branched poly(lactide)s (PLAs) and PLGs with low molecular multifunctional alcohols, like glycerol, pentaerythritol, mannitol/sorbitol or star-shaped poly(ethylene glycol)s, as described e.g. in: S. H. Kim et al.: Preparation of star-shaped polylactide with pentaerythritol and stannous octoate; Makromol. Chem., vol. 194 (1993), pages 3229-3236;        using comb-like, highly branched polyesters synthesized by grafting sulfonic modified dextran based, backbones with lactide and glycolide, which allow a faster biological degradation with negatively charged groups, as described e.g. in: Li et al.: Biodegradable brush-like graft polymers from poly(D,L-lactide) or poly(D,L-lactide-co-glycolide) and charge-modified, hydrophilic dextrans as backbone-Synthesis, characterization and in vitro degradation properties; Polymer 1997; vol. 38, pages 6197-6206;        the synthesis of biodegradable comb PLGA by grafting short PLGA chains onto different poly(vinyl alcohol) (PVA) based backbone polyols, poly(2-sulfobutyl vinyl alcohol) and poly(diethylaminoethyl-vinyl alcohol). In this system, the adjustment of the polymer-properties were carried out by introducing charged groups sulfobutyl moieties or amine structures into the PVA backbone, to create polymers with negative or positive charges, as described e.g. in: Breitenbach A, Kissel T.: Biodegradable comb polyester: Part 1. Synthesis, characterization and structural analysis of poly(lactide) and poly(lactide-co-glycolide) grafted onto water-soluble poly(vinyl alcohol) as backbone; Polymer, 1998; vol. 39: pages 3261-3271.        
By another approach to effectively load positively charged drugs, proteins and peptides, functional groups were introduced to the drug delivery systems, such as hydroxycarboxylic groups, as disclosed e.g. in WO 95/23175 A1.
Also through the NaH activation method the sulfobutylated PVA were obtained and using them to graft PLGA, the negatively charged poly(2-sulfobutyl vinyl alcohol)-g-PLGA were prepared, leading to a better temperature stability during bulk polymerization with PLGA (Breitenbach A. et al.: Biodegradable comb polyesters containing polyelectrolyte backbones facilitate the preparation of nanoparticles with defined surface structure and bioadhesive properties; Polym. Adv. Technol.; 2002; vol. 13, pages 938-950).
Because of the NaH activation method the synthesis of the negatively charged poly(2-sulfobutyl vinyl alcohol)-g-PLGA is difficult to control with respect to the degree of substitution which can be reached through the introduction of the sulfobutyl groups to the PVA backbone.
Additionally, the sulfobutylation approach does provide only a small range of variations of the degree of substitution with negative charges.
As can be seen from these explanations there still exists a need for further polymers, which are better suitable as appropriate carriers for biologically or pharmacologically active substances and which allow effective loading and sustained-release of especially positively charged drugs, proteins and peptides from micro- or nanospheres or micro- or nanoparticles as polymeric drug delivery systems. It must be kept in mind that these pharmacologically active substances make up a quite diverse group of molecular entities, characterized by different physicochemical properties and different requirements with regards to the application.
It is therefore an aim of the present invention to provide a set of novel polymers which could be used as polymeric drug delivery systems (polymeric excipients) for at least one, preferably more classes of pharmaceutically active ingredients.
It is an additional task to provide polymers which are especially well suited as carriers for biologically and/or pharmacologically active molecules like proteins and peptides and drugs, especially those which are cationic and/or positively charged. The overall biophysical properties of such materials had to be specifically designed to allow modifications of the drug loading and degradation behaviour of the desired drug delivery system.
It is a further task to provide polymers which allow the preparation of micro- or nanoparticles and of self-aggregating colloidal systems with defined and finely adjustable surface properties. Preferably this preparation of micro- or nanoparticulate systems should be possible even without the use of surfactants in the water phase during their preparation, thus qualifying the resulting polymers for the use in the respiratory tract.
As a solution to these problems the present invention provides graft copolymers of poly(vinyl sulfonic-co-vinyl alcohol)-g-poly(lactide-co-glycolide) (P(VS-VA)-g-PLGA) with negatively charged electrolyte properties.
Graft copolymers according to the invention provide novel polymeric excipients with hitherto unknown properties which make them well suited as drug delivery systems. These systems allow effective loading and sustained-release of drugs, proteins and peptides, especially for medical applications. They can also be applied to the preparation of drug-loaded particles, microparticles and nanoparticles and self-aggregating colloidal systems. Drug delivery systems based on this material are especially useful as parenteral or mucosal drug delivery systems for pharmaceutical applications.
The amphiphilic and negatively charged nature of the graft copolymers according to the invention allows a better drug loading by electrostatic interactions as a function of sulfonic substitution. They are suitable for generally all pharmacologically active substances and for effective loading and sustained-release of such active substances, especially of positively charged drugs, proteins and peptides.
The particular advantage of this polymer type is the broad range of the degree of substitution with negative charges which cannot be achieved by e.g. sulfobutylation.
Thus it is an advantage of graft copolymers according to the invention that they can reproducibly be synthesized with even minute variations of the chemical structure. For example, the degree of sulfonic substitution and the degree of PLGA substitution as well as composition of the PLGA side chains can be designed. This also allows the preparation of micro- and nanoparticles and self-aggregating colloidal systems with accurately defined properties such as inertness against drugs or biopolymers like proteins or nucleic acids, intrinsic viscosity, solubility, melting point, glass transition temperature, hydrophilicity/hydrophobicity (amphiphilic nature), rate of degradation and surface charge. Especially the substitution degree of negative charges can be varied over a wide range, which is wider e.g. than the respective variation degree of sulfobutyl-polyvinylalcohol—graft PLGA (SB-PVAL-g-PLGA).
For example a system according to the invention can be designed for the loading with positively charged drugs like Salbutamol, Ipratropiumbromid or Tiotropiumbromid or others (see below). The charge allows a slower release of the drug in vivo in comparison to a more neutral material.
Additionally, in most cases the preparation of micro- and nanoparticles and self-aggregating colloidal systems from these polymers is possible even without the use of surfactants in the water phase.
These polymers combine, within a modified three-dimensional architecture, increased hydrophilicity (due to the grafted side chains) and negatively charged groups (due to the vinyl sulfonic) in a single polymer, all of which can be precisely synthesized.
Due to the existence of charged and/or polar groups, graft copolymers according to the invention can be mixed with a higher amount of drugs with positively charged groups than neutral carriers can. Under application conditions an active drug is released more slowly by a material according to the invention than by a material without anionic groups.
Aiming at a pharmaceutical use of graft copolymers according to the invention, it should be noted that they are largely bio-degradable because polymers from hydroxycarboxylic acids like lactic and/or glycolic acid are hydrolyzed in the patient's body to lactic and/or glycolic acid, which are further metabolized to CO2 and water. Graft copolymers according to the invention are therefore particularly advantageous for the production of parenterally applicable preparations. Variations in the structure can be used to modify the rate of degradation of the graft polymers according to this invention.
Graft copolymers according to the invention can be applied in all industrial areas that refer to polymers. They are especially useful as components of drug delivery systems allowing formulations of pharmaceutically active ingredients, especially for providing slow-release forms of drug delivery systems which can be used e.g. for parenteral or mucosal applications.
Graft copolymers according to the invention exhibit a core-corona structure with the negatively charged hydrophilic sulfonic groups oriented towards the outer aqueous phase which provides an optimal surface for the loading of cationic substances. This structure allows the preparation of colloidal carriers without the use of additional surfactants. This is extremely important for pulmonary applications, as the inhalation of significant amounts of surfactant may disturb the naturally regulated surface tension of the pulmonary lining fluid, thus leading to impaired lung function or inflammation.